Hi, I have started my PhD in Lisbon and planning to create mABs using phage display technique. I did some research and didn't understood how can I create my library, of course I can go for the commercial, but still want to have my own. Did anyone have some info/protocol and maybe suggestions about costs of doing that?
See:
Schaefer, J. V., Honegger, A. and Plückthun, A. (2010) Construction of scFv fragments from hybridoma or spleen cells by PCR assembly in: Antibody Engineering 2nd ed. (Kontermann, R., and Dübel, S., eds) Vol. 1 pp. 21-44. Springer Verlag, Berlin, Heidelberg, Germany
https://plueckthun.bioc.uzh.ch/wp-content/uploads/Publications/APpub0307.pdf
based on and improved from:
Burmester, J. and Plückthun, A. (2001) Construction of scFv fragments from hybridoma or spleen cells by PCR assembly in: Antibody Engineering(Kontermann, R., and Dübel, S., eds) pp. 19-40. Springer-Verlag, Berlin, Heidelberg
https://plueckthun.bioc.uzh.ch/wp-content/uploads/Publications/APpub0208.pdf
Krebber, A., Bornhauser, S., Burmester, J., Honegger, A., Willuda, J., Bosshard, H. R. and Plückthun, A. (1997) Reliable cloning of functional antibody variable domains from hybridomas and spleen cell repertoires employing a reengineered phage display system. J. Immunol. Methods 201, 35-55
https://plueckthun.bioc.uzh.ch/wp-content/uploads/Publications/APpub0113.pdf
If interested, contact the Plückthun lab for the vectors used in the study. You would probably have to have the oligonucleotides synthesised, as a good quality is essential and you would not want to rely on stuff that has lingered in a freezer for a decade. For the reagents, consult the appropriate catalogs. The costs for equipment very much depends on what is already available in the lab where you are working.
Building an antibody library is easier said than done. What kind of antibody library, with the natural diversity from donors, synthetic? semisynthetic? You can find an example of a library built from a single donor in this manuscript, but there are really a lot available online:
https://www.nature.com/articles/s42003-021-01881-0
Quality control, i.e. the assessment of diversity, is essential. If possible, resort to something that is available and tested. Will save you time, money, resources and frustration. Not sure if the materials still are available (couldn't find it by a quick google search), but I'd like to recommend checking out the Tomlinson I+J ScFv library, which hopefully still is available for academic use at a very reasonable cost.
For details of its application, see e.g.
Article Generation of Novel Single-Chain Antibodies by Phage-Display...
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